This project is aimed at developing and demonstrating the performance of a new multi-stage time-of-flight mass spectrometer that provides accurate measurement of the relative abundance of isotopes at very low levels. This instrument will ultimately provide performance at least equal to that achieved by established methods employing accelerator mass spectrometers at a very small fraction of the cost. The proposed bench top instrument is small, highly automated and suitable for use by relatively untrained operators in a hospital or medical research laboratory. The major focus in this work is on those applications that require measurements of specific isotopes at levels below the part-per-billion level. These applications generally involve radioactive isotopes with very long half-lives (>1000 years). Specific examples include 14C for radiocarbon dating and biological tracer studies, and 41Ca used as a tracer for monitoring bone long term metabolic status in human patients. These applications often require precise determination of the relative abundance of isotopes at levels below 10-10 and extending down to 10-15. At present these measurements require use of a very large and expensive accelerator mass spectrometer (AMS) generally located in a central facility. Accelerator mass spectrometry has demonstrated the utility of long-lived radioisotopes as biological tracers, but applications have been limited by the relatively hig cost and inaccessibility of complex instruments housed in central facilities. Nevertheless, support from the NIH, the pharmaceutical industry, and several AMS-based businesses led to a 2006 FDA guidance statement including AMS-based 14C micro dosing (i.e., first-in-human Phase 0 studies) of drug pharmacokinetics and pharmacodynamics, and AMS studies of 41Ca for cancer diagnostics have also seen early support from the NIH. The technique is limited to solid samples deposited on a suitable target, and it appears that the sample preparation procedures that have been developed for AMS can be employed with little or no modification in the proposed instrument. In addition to developing an economical alternative to AMS for measuring radioactive isotopes at ppt levels, this project will provide a very powerful method for accurate measurement of more abundant isotopes such as 2H, 13C, 15N, 17O, 18O, 33S, and 34S, as well as a wide variety of metals even when present in very complex matrices. SIMS has already been demonstrated for applications to biological tissues, microorganisms, and other complex organic and inorganic samples using conventional SIMS instrumentation. The high mass resolving power, high speed, and high sensitivity available with the new multi-stage TOF analyzer should provide superior results for these applications, and will allow simultaneous accurate measurements of stable isotope levels at sub-picomole levels for components separated by LC. An approach to identification of components in LC fractions by tandem TOF MS in parallel with simultaneous measurements of isotope ratios is presented. Two major innovations in time of flight mass spectrometry make this project feasible. One is the development of a multi-stage TOF mass spectrometer that dramatically improves the abundance sensitivity compared to earlier instruments. The other is invention and implementation of a new principle in time of flight mass spectrometry that provides simultaneous space and velocity focusing with either pulsed or continuous sources of ionization. The latter enables the use of a Cs+ SIMS ion source that provides several orders of magnitude greater ionization rates that are possible with the pulsed lasers used in earlier work. PUBLIC HEALTH RELEVANCE: The introduction of biomedical applications of AMS in the early 1990's brought the possibility of significant changes in strategies employed for drug development. The ability of AMS to distinguish 14C-labeled compounds from their unlabeled counterparts and to quantify exceedingly small amounts of these compounds in any biological matrix enabled micro dosing studies to obtain detailed information on metabolic pathways long before safety and efficacy trials in humans. Measurement of 41Ca/Ca in urine and serum enables early detection and improved clinical management of osteoporosis, multiple myeloma, and cancer metastatic to the bone. This project will enable broader application of these state-of-the-art research and diagnostic methods by providing inexpensive instruments with competitive performance that are suitable for routine use in clinical and medical research laboratories.